Paramyxoviruses are the leading causative agents of acute viral respiratory tract infections. Among the paramyxoviruses, human metapneumovirus (hMPV), human respiratory syncytial virus (RSV), and human parainfluenza virus type 3 (hPIV3) account for more than 70% of acute viral respiratory diseases. All of three viruses cause similar clinical signs and symptoms, ranging from mild respiratory problems to sever coughs, bronchiolitis, and pneumonia. All three viruses cause acute respiratory tract disease in individuals of all ages, especially in infants, children, the elderly, and immunocompromised individuals. In the United States, 60% of infants are infected during their first RSV season, and nearly all children will have been infected with the virus by 2-3 years of age. HMPV is a newly discovered human pathogen, first identified in 2001 in The Netherlands. Soon after its discovery, hMPV was recognized as a globally prevalent pathogen. Epidemiological studies suggest that 5 to 15% of all respiratory tract infections in infants and young children are caused by hMPV, a proportion second only to that of RSV. PIV3 is the third causative agent of viral respiratory infection in children and infants. All three pathogens are globally prevalent.
Despite the enormous economic losses and emotional burdens these viruses cause, vaccines and anti-viral drugs are currently not available. For decades, approaches to generate vaccines employing viral proteins or inactivated vaccines have failed either due to a lack of immunogenicity or the potential for causing enhanced pulmonary disease upon natural infection with the same virus.
Vaccination has been the most effective public health strategy to reduce morbidity and mortality associated with viral infections. The increasing clinical significance of RSV, hMPV and PIV3 infections suggest that there is an urgent need for a safe and efficacious vaccine against these viruses, particularly for the populations at high risk such as infants, children, elderly, and immunocompromised individuals. An effective vaccine would not only prevent acute respiratory tract infection caused by these viruses, but also block transmission routes and thus improve human and public health. In the current inventions, the inventors developed a panel of live attenuated vaccines against RSV, hMPV and PIV3.
However, development of vaccine against human paramyxoviruses has met serious challenge. With the exception of the influenza virus, there is no FDA approved vaccine for other viruses that cause acute upper and lower respiratory tract infections in human. Generally, inactivated and live attenuated vaccines are the two most common strategies used in vaccines against infectious diseases. For safety, an inactivated vaccine is preferred. However, development of an inactivated vaccine for the paramyxovirus RSV turned out to be a problem. A formalin-inactivated RSV vaccine developed and tested in the 1960s not only failed to induce a protective immune response in human, but led to an enhanced respiratory disease upon natural infection with RSV. Eighty percent of the vaccinated children were hospitalized following natural RSV infection, and two children died. Enhanced respiratory disease following vaccination with inactivated vaccine has been observed in other paramyxoviruses such as PIV-3, hMPV and measles virus. A recent study showed that cotton rats immunized with inactivated hMPV vaccine were protected against infection, but developed increased lung pathology. These observations suggest that inactivated vaccines are not the primary choices for three paramyxoviruses: hMPV, RSV, and PIV3, all of which cause extensive morbidity and mortality in the same population, infants and children.
In contrast to inactivated vaccines, enhanced lung diseases have not been observed for candidate live attenuated RSV vaccines. Therefore, live attenuated vaccines are the most promising vaccine candidates for hMPV, RSV, and PIV3. However, it has been technically challenging to isolate a virus with low virulence that retains high immunogenicity. In paramyxoviruses, spike proteins (F and G proteins for RSV and hMPV) are major determinants of virulence. Therefore, traditional attenuation strategies have been focused on engineering mutations in these two glycoproteins. However, F and G proteins are also viral immunogenic antigens that are responsible for immune response. As a consequence, mutations in glycoproteins may impair the immunogenicity of the attenuated live vaccine. Therefore, exploration of new attenuation approaches is urgently needed.
This invention develops new attenuated viruses as live vaccine candidates for major human paramyxoviruses including hMPV, RSV and PIV3 by targeting viral mRNA cap methyltransferase (MTase). Paramyxoviruses share a common strategy for replication and gene expression. During RNA synthesis, paramyxoviruses yield capped, methylated, and polyadenylated mRNAs. Methylation of the mRNA directly impacts the stability of mRNA and subsequent translation of viral proteins, which in turn affects viral genome replication, virus assembly, and budding. The large (L) polymerase protein catalyzes the mRNA cap MTases. Recombinant virus defective in MTase can be recovered from cloned full-length viral cDNA by a reverse genetics system. Viruses lacking MTase would likely be attenuated without affecting immunogenicity, since the MTase is located in L protein, which is not a neutralizing antibody target. Thus, ablating viral mRNA cap methylation provides a new avenue to rationally attenuate these viruses for development of live attenuated vaccines.